The polymerase chain reaction (PCR) is a powerful method for amplifying nucleic acid sequences. Various disclosures involving this technique are found in U.S. Pat. Nos. 4,683,202; 4,683,195; 4,800,159; 4,965,188; and 5,512,462, each of which is incorporated herein by reference. In a simple form, PCR is an in vitro technique for the enzymatic synthesis of specific DNA sequences using two oligonucleotide primers that hybridize to complementary nucleic acid strands and flank a region that is to be amplified in a target DNA. A series of reaction steps of 1) template denaturation, 2) primer annealing, and 3) extension of annealed primers by DNA polymerase, results in the geometric accumulation of a specific fragment whose termini are defined by the 5′ ends of the primers. As is well known, PCR is capable of selective enrichment of specific DNA sequences by a factor of 109.
PCR has been applied widely in molecular biology for sequencing, genome mapping and forensics. However, despite such wide-spread use, amplifying long stretches of DNA, particularly genomic DNA, is difficult. Many protocols for long range PCR exist; however, reaction conditions are usually optimized for amplifying specific target regions of interest. Applying the same “optimized” reaction conditions to amplify a different target region may not result in a detectable amplification product.
In light of the above limitations, there is a need in the art for methods capable of amplifying nucleic acid sequences. The resulting methods may be used in some embodiments to amplify mammalian target sequences across the genome to facilitate genotyping studies, and for other applications in the art of molecular biology.